The project described in this application addresses a mechanistic study aimed at understanding the steps involved in generation of immunodominant epitopes. Immunodominance is a phenomenon that has long been recognized but yet remains unclear to date. It is well known that the immune system focuses on and responds to very few representative epitopes (referred to as immunodominant epitopes) from invading pathogenic insults ranging from such as infectious agents and, antigenic targets in autoimmune diseases, allergy, and cancer. In each all these cases, the immune system either responds positively or fails to respond to antigenic peptides in the context of MHC molecules. Recent advances in our understanding of the antigen presentation pathway have shown that the steps of antigen processing and selection critically influence the peptide repertoire presented to T-cells. Recently, we have made considerable progress in developing a reductionist antigen processing system for MHC class II molecules that utilizes five purified protein components of the class II antigen presentation pathway. Notably, this system yielded physiologically relevant immunodominant epitopes restricted to HLA-DR1. In this proposal, we will extend this MHC II system to another MHC class II molecules HLA-DR4 and would explore steps involved in epitope capture and antigen processing. Overall, we propose to dissect underlying factors of T-cell immunodominance. Aim 1 would explore the temporal relationship between antigen capture and processing. In Aim 2, we would investigate epitope hierarchy and the role for DM, and in Aim 3, we would analyze contributions of HLA-DO to epitope capture and editing. Hence, knowledge of the identity of immunodominant epitopes, and a clear understanding of how they are generated inside cells, can guide the design of effective immunotherapeutics.